The compatible solute transporters of Listeria monocytogenes: Their roles in hyperosmotic and chill tolerance

The food-borne human and animal pathogen Listeria monocytogenes is known for its tolerance to conditions of elevated osmotic strength and low temperature, both of which are directly associated to the organism's ability to accumulate compatible solutes (osmolytes) from its environment.; Accumulation of osmolytes was investigated in cytoplasmic extracts of L. monocytogenes grown in bovine milk whey at 7 and 30°C. In balanced growth cultures at 7°C, glycine betaine and carnitine were the major osmolytes accumulated by wild type L. monocytogenes 10403S, whereas carnitine was the dominant osmolyte in LTG59, a mutant blocked in the major glycine betaine uptake system, glycine betaine porter II. At 4°C strain LTG59 grew with a longer lag phase and generation time and reached a lower final cell yield than that of strain 10403S.; A Tn917-LTV3 insertional mutant (LTS4a) was isolated that could not be rescued from hyperosmotic stress by exogenous carnitine. LTS4a grew indistinguishably from a control strain (DP-L1044) in the absence of stress or in the absence of carnitine, but DP-L1044 grew substantially faster under osmotic or chill stress in the presence of carnitine. LTS4a was found to be strongly impaired in KCl-activated as well as chill-activated carnitine transport. Direct sequencing of LTS4a genomic DNA allowed us to determine that the opuC operon had been interrupted by the transposon. opuC encodes an ABC carnitine transporter that can be activated by either hyperosmotic stress or chill and plays a significant role in the tolerance of L. monocytogenes to both forms of environmental stress.; The specificity of the two glycine betaine porters and OpuC towards glycine betaine and carnitine was investigated by creating mutant derivatives of L. monocytogenes 10403S that possess each of the transporters in isolation. Kinetic and steady state osmolyte accumulation data together with growth rate experiments demonstrated that osmotically activated glycine betaine transport is readily and effectively mediated by Gbu and BetL but only weakly by OpuC. Osmotically stimulated carnitine transport was demonstrated for OpuC and Gbu. No other transporter in L. monocytogenes 10403S appears to be involved in transport of either osmolyte since a triple mutant strain yielded neither transport nor accumulation of glycine betaine or carnitine and could not be rescued by either osmolyte when grown under elevated osmotic stress.; The inclusion of osmolyte content of foods as variables in bacterial growth modeling studies and HACCP programs will enhance their accuracy and predictive value and therefore contribute to food safety and prevention of food spoilage.